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为了解ampD ampR调节基因与阴沟肠杆菌AmpC酶表达的关系 ,对 58株阴沟肠杆菌采用表型筛选法初筛 ,PCR法扩增AmpC酶的调节基因ampD和ampR ,并对部分PCR产物进行克隆测序。表型筛选结果显示 :去阻遏高产突变株为 50株 ,高度诱导产酶株为 3株。PCR法扩增目的基因片段显示 ,4 9株携带ampD基因 ,51株携带ampR基因。对其中 15株细菌的ampD基因和ampR基因进行克隆测序 ,发现 3株高度诱导型中 2株ampD基因存在 95位氨基酸的突变位点 ,而ampR基因未发现突变位点 ;12株去阻遏高产型中 ,8株ampD基因存在羧基端可疑的突变位点 ,5株ampR基因存在可疑的突变位点。结果提示 :ampD蛋白羧基端的氨基酸缺失或替代可能与AmpC酶的去阻遏表达有关 ,ampR基因的突变位点可能影响AmpC酶的去阻遏表达
To understand the relationship between ampD ampR regulatory gene and Enterobacter cloacae AmpC enzyme expression, 58 strains of Enterobacter cloacae were screened by phenotypic screening. AmpC and ampR were amplified by PCR, and some PCR products were cloned Sequencing. Phenotypic screening results showed that 50 strains of high-yielding mutants were repressed and 3 strains of highly-induced enzyme-producing strains. PCR amplification of the target gene fragment showed that 49 strains carried ampD gene and 51 strains carried ampR gene. Among them, ampD gene and ampR gene of 15 strains of bacteria were cloned and sequenced. It was found that there were 95 amino acid mutation sites in ampD gene among the three highly inducible genes, but no mutation was found in ampR gene. Among the 8 strains of ampD gene, there were some suspicious mutations in the carboxyl terminal region and 5 strains of ampR gene had a suspicious mutation site. The results suggest that the amino acid deletion or substitution at the carboxyl end of ampD protein may be related to the de-repression expression of AmpC enzyme. The mutation site of ampR gene may affect the expression of AmpC enzyme